The long-term goal is to find how axons in the vertebrate brain determine when and where to form synapses. The current focus is on a fundamental determinant of connectivity throughout the brain, whereby axonal populations confine their terminal arbors and synapses to specific laminae within a target area. The experimental preparation is retinotectal projection in the chick, because it is experimentally-accessible and exquisitely laminated. Moreover, all retinal axons terminate in just 3 of 16 tectal laminae, the retinorecipient laminae (RRL), and each retinal axon terminates in just one of the RLL. Initial studies identified a set of five adhesive macromolecules that are selectively expressed in the RRL and showed that three of them, N-cadherin, SC1/DM-GRASP and a glycoprotein recognized by the Vicia villosa B4 agglutinin lectin (VVA-B4), are essential for lamina-specific arborization of retinal axons. To extend this work, aim one will identify the glycoprotein that binds VVA-B4 and elucidate the mechanisms that localize N-cadherin and SC1 to the RRL. Aim 2 will assess the roles of these three molecules in synapse formation per se, using both tissue slices and cultures of dissociated retinal and tectal cells. Aim 3 will test the hypothesis that, just as N-cadherin promotes the laminar selectivity of retinal axons, other cadherins promote lamina-specific behaviors of other tectal inputs. To this end, preliminary results have identified partial sequences of 24 distinct cadherins expressed in tecta. Aim 4 will test the hypothesis that one set of molecules (including N-cadherin and SC1) is involved in targeting retinal axons to RRL in general, whereas a second set promotes arborization of axonal subsets in individual laminae. To this end, this aim will examine selective innervation of individual retinorecipient laminae by defined subsets of retinal ganglion cells. Aim 5 will begin to extend analysis from the experimentally-accessible chick retinotectal synapse to the genetically accessible mouse retinocollicular synapse by completing the development of a method for selectively expressing neuronal genes along with an axonal marker in retinal ganglion cells of transgenic mice. The hope is that this strategy will eventually elucidate mechanisms that promote laminar-specific synapse formation in mammals.